Development and Evaluation of Novel Aptamer-based Therapeutics Targeting SARS-CoV-2 in a Physiologically-Relevant Model of Human Airway Epithelium

在人类气道上皮生理相关模型中针对 SARS-CoV-2 的新型适体疗法的开发和评估

• 批准号: 10287842
• 负责人: JEFFREY J DESTEFANO
• 金额: $ 23.18万
• 依托单位: UNIV OF MARYLAND, COLLEGE PARK
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-12 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10287842
• 关键词: 2019-nCoV; ACE2; Affinity; Alveolar; Antibodies; Antiviral Agents; Aptamer Technology; Base Sequence; Binding; Biochemical; Biological Assay; Biosensor; COVID-19; COVID-19 therapeutics; COVID-19 treatment; COVID-19 vaccine; Cell Culture Techniques; Cells; Cessation of life; Chemical Structure; Clinical; Collaborations; Containment; Coronavirus; DNA; Development; Diagnostic; Disease; Drug Screening; Enzyme-Linked Immunosorbent Assay; Evaluation; Evolution; FDA Emergency Use Authorization; FDA approved; Fatality rate; Feedback; Future; Generations; Goals; Human; In Vitro; Infection; Infection prevention; Letters; Ligands; Link; Membrane Proteins; Middle East Respiratory Syndrome; Modeling; Modernization; Modification; Molecular Biology; Morbidity - disease rate; Mucous Membrane; Nucleic Acids; Nucleotides; Outcome; Pharmacotherapy; Phase III Clinical Trials; Physiological; Prevention; Property; Proteins; Public Health; RNA; Race; Resistance; Respiratory Failure; Respiratory System; Ribonucleases; SARS coronavirus; Societies; Surface; System; Testing; Therapeutic; Toxic effect; United States; Upper respiratory tract; Vaccines; Vertebral column; Viral; Viral Pneumonia; Virus; Virus Diseases; Virus Inhibitors; Work; airway epithelium; alveolar type II cell; aptamer; base; human model; human pathogen; immunogenicity; improved; in vitro Model; in vivo; innovation; inorganic phosphate; mortality; new therapeutic target; novel; novel therapeutics; nucleotide analog; pandemic disease; particle; rapid test; receptor binding; remdesivir; respiratory virus; screening; small molecule; sugar; technology development; therapeutic target; tool; uptake; vaccine candidate; virus host interaction

The impact of SARS-CoV-2 on public health and the global economy cannot be overstated. As of September 28, 2020, 33,224,222 cases and 999,298 deaths worldwide have been linked to this emergent virus. This staggering number continues to grow, with the United States baring disproportionately high rates of morbidity and mortality. The virus targets the respiratory tract, leading to a wide range of clinical outcomes including mild upper respiratory tract illness and severe viral pneumonia with respiratory failure. To date, four SARS-CoV-2 vaccine candidates have entered phase 3 clinical trials and a massive parallel effort has been undertaken to repurpose already FDA-approved drugs for the treatment of COVID-19 or identify compounds with potential therapeutic activity. Despite this effort, remdesivir remains the only approved (with emergency use authorization) direct-acting antiviral for the treatment of COVID-19. Of critical importance: there is currently no vaccine or SARS-CoV-2-specific therapy approved for the prevention or treatment of disease. Furthermore, multiple antivirals may be required to avoid the rapid emergence of resistant SARS-CoV-2 strains. Thus, the development of novel therapeutics targeting SARS-CoV-2 are urgently needed. Infection requires interaction between the viral surface protein, spike (S), and a host protein, ACE2, that is expressed on type II alveolar cells and ciliated cells in the human airway epithelium (HAE), making these cells potentially vulnerable to infection. Thus, our goal is to develop a novel therapeutic that blocks this interaction between spike (on the virus) and ACE2 (on the host cell) to prevent infection and ameliorate disease. Aptamers are short nucleic acid-based sequences that bind with high affinity to their targets. Among other applications, aptamers have been shown to have potent antiviral activity and low toxicity in cell culture. While aptamers were originally made with RNA and DNA, Xeno-Nucleic Acids (XNA: nucleotide analogs with altered sugar, base, or phosphate backbones) have emerged as important new substrates and XNA aptamers often demonstrate enhanced target binding and greater stability compared to RNA and DNA aptamers. Thus, we hypothesize that aptamer technology, and specifically XNA aptamers, can be leveraged to inhibit spike-ACE2 interaction and propose to establish an innovative, in vitro screening platform that can serve to assess the efficacy of such aptamers, or other novel therapeutics, in blocking infection. This platform will utilize SARS-CoV-2 pseudoparticles (allowing work under Biosafety Level 2 containment) and a physiologically-relevant in vitro model of human airway epithelium that recapitulates the mucosal surface of the airway in vivo. Aptamers will also be tested using live virus infections of culture cells (Biosafety Level 3). This work is highly significant given the immediate need for novel therapeutics against SARS-CoV-2. Further, the development of a high-throughput, pseudoparticle-based assay to assess viral entry in a relevant culture system will have broad applications for additional drug screens and / or studies that aim to further understand SARSCoV- 2 virus-host interactions at the level of particle uptake.

SARS-COV-2对公共卫生和全球经济的影响不能被夸大。截至9月 28，2020，33,224,222例和999,298例死亡与这种新兴病毒有关。这 惊人的数字继续增长，美国的发病率不成比例 和死亡率。该病毒针对呼吸道，导致广泛的临床结局，包括轻度 上呼吸道疾病和严重的病毒性肺炎，呼吸衰竭。迄今为止，四个SARS-COV-2 候选疫苗已经进入了第三阶段临床试验，并且已经采取了巨大的平行努力 重新利用已经由FDA批准的药物用于治疗COVID-19或鉴定具有潜在的化合物 治疗活动。尽管做出了这项努力，雷姆迪维尔仍然是唯一的批准（紧急使用授权） 直接作用抗病毒治疗COVID-19。至关重要的是：目前没有疫苗或 SARS-COV-2特异性治疗已批准用于预防或治疗疾病。此外，多个 可能需要抗病毒药以避免抗性SARS-COV-2菌株的快速出现。因此，发展 迫切需要针对SARS-COV-2的新型治疗剂。感染需要病毒之间的相互作用 表面蛋白，尖峰（S）和宿主蛋白ACE2，在II型肺泡细胞和纤毛细胞上表达 在人类气道上皮（HAE）中，使这些细胞可能容易受到感染的影响。因此，我们的目标是 开发一种新型的治疗性，可以阻止尖峰（病毒）和ACE2（在宿主细胞上）之间的相互作用 防止感染和改善疾病。适体是基于核酸的短序列，与 对目标的高亲和力。除其他应用外，适体已显示具有有效的抗病毒活性 和细胞培养中的低毒性。最初用RNA和DNA制成的适体，而异种核酸 （XNA：用改变糖，碱或磷酸盐骨架改变的核苷酸类似物）已出现为重要的新 与RNA相比 和DNA适体。因此，我们假设适体技术，尤其是XNA适体可以是 利用以抑制Spike-ACE2相互作用并提议建立创新的体外筛选平台 可以评估这种适体或其他新型治疗剂在阻断感染中的功效。这 平台将利用SARS-COV-2伪粒子（允许在生物安全2级遏制下进行工作）和 人类气道上皮的体外模型与生理上的体外模型，该模型概括了粘膜表面 气道在体内。适体还将使用培养细胞的活病毒感染（生物安全3级）进行测试。这 鉴于对SARS-COV-2的新型治疗方法的立即需要，工作非常重要。此外， 开发高通量，基于假颗粒的测定法，以评估相关培养系统中的病毒入口 将在其他药物筛查和 /或旨在进一步了解Sarscov-的其他药物筛查和 /或研究中广泛应用 2在颗粒吸收水平上的病毒宿主相互作用。